In the search for alternative agricultural insectides which will control insect pests with minimal environment impact, attention is now turning to biological agents for such control. Entomopathogenic nematode families which include Steinernematidae and Heterorhabditidae offer such a possibility, being natural pathogens of insects.
Entomopathogenic nematodes are found in soil as the third stage "infective" juvenile, which is able to seek out and penetrate insect larvae/adults. These nematodes carry specific symbiotic bacteria Xenorhabdus nematophilus/luminescens which assist in the killing of the insect. Once inside the insect, the nematodes interfere with the insect's immune system and release symbiotic bacteria which multiply rapidly and breakdown the haemocoel (insect blood/tissue). The nematodes begin to feed on the bacteria and the partially digested haemocoel, mature and reproduce. Together the nematodes and the bacteria cause death of the insect within 24-72 hours.
The concept of pest control with entomopathogenic nematodes is based on mass rearing of nematodes and effective application to insect pests susceptible to nematodes. Progress in exploitation of entomopathogenic nematodes for biological control has been hindered due to problems mainly with mass rearing and the inability to efficiently store them. In the past rearing of nematodes was restricted to in vivo methods such as on Galleria mellonella, but recently much progress has been made with in vitro rearing. Now that artificial mass rearing has been achieved, the feasibility of using entomopathogenic nematodes as commercial bioinsecticides is more realistic.
Storage and transport of nematodes have emerged as the crucial factors before nematodes are accepted as a practical alternative to chemical insecticides.
It is known that the third stage juvenile can survive in soil for an extended period and under very extreme conditions such as desiccation and freezing, revive as the condition becomes favourable and infect insects when they emerge, however the numbers surviving these conditions are not large.
The nematode is, on the other hand, particularly vulnerable to quick desiccation and exposure to UV radiation and high temperature.
The third stage infective juvenile, a non-feeding stage, lives on stored substrates.
It is likely that the survival capacity of a given nematode species will relate to its ability to accummulate particular energy reserves, such as glycogen or lipids, during the growth phases prior to the desired infective stage, and to mobilise these at a rate sufficient to meet non-growth (survival) demands. Excessive mobilisation in environments not conducive to infection of the target insect host by this infective third-stage would constitute wasteful energy consumption and reduce duration of survival and subsequent chance of infection. Thus survival time may in part be determined by the available energy store, but may also involve several additional factors such as, enzyme deactivation and the generation and excretion of toxic metabolic by-products.
Nematodes, currently used for efficacy tests on certain insects, are stored at a reduced temperature, 1.degree.-10.degree. C., where the rate of catabolism in the nematode is reduced.
Oxygen is either fed continuously/intermittently or provided by limiting the number of nematodes in sealed containers. Many workers have emphasised importance of oxygenation and large surface area-to-volume ratio for oxygen exchamge when storing nematodes (Dutky et al, 1964 and Hara et al 1981). The contact area of the nematode suspension to air is increased by using filter paper, tracing paper, cottonwool, sponge etc. For example, 500 million Steinernema feltiae and S. bibionis can be stored on 100 g clean dry sponge for up to 4 months in a continuously aerated bag at 1.degree.-2.degree. C. (Bedding, 1984).
However, the above workers discuss the preservation of nematodes in the context of supplying nematodes for laboratory and field trials.
Generally the methods for storing nematodes described for field trials and laboratory use are not suitable for a wide commercial scale distribution due to shortcomings such as:
(i) requirement of refrigeration; PA1 (ii) requirement of oxygen (provided through aeration/ample air space); PA1 (iii) limited nematode density (up to 150,000/ml) even with abovementioned provisions; and PA1 (iv) difficulty to extract nematodes if any carrier is used.